The plant is reported to have anti-inflammatory and antiarthritic properties. [Wealth of India: Raw Materials, CSIR, New Delhi, 1976, vol. X, p. 522]. A variety of constituents have been reported from this plant. From the leaves Ghosh and Krishna isolated glucononitol, p-hydroxybenzoic acid, 5-hydroxyisophthalic acid and 3,4-dihydroxy-benzoic acid along with two glucosides and an amorphous alkaloid [Ghosh, T. P. and Krishna, S., Indian Chem. 1936, 13, 634] In the essential oil of leaves a-pinene, camphene, citral and (3-caryophyllene have been reported [Masilungan, V. A., Philip. J. Sci, 1955, 84, 275].
A large number of flavonoids viz., casticin, orientin, isoorientin, luteolin, luteolin-7-0-glucoside, corymbosin, gardenins A and B, 3-0-desmethylartemetin,5-0-desmethylnobiletin,3\4\5,5\6J,8-heptamethoxyflavanone and 3\,5-dihydroxy-4′,6,7-trimethoxy flavanone and 3′,5-dihydroxy-4′,6,7-trimethoxyflavanone have been reported from this plant [Sirait, L. M., Rimpler, H. and Haensal, R., Experientia 1962, 18, 72; Haensal, R. et al. Phytochemistry 1965, 4 ,19; Banerji A. et al. Phytochemistry 1969, 8,511; Ferdous, A. J. et al. Bangladesh Acad. Sci. 1984, 8,23; Dayrit, F. M. et al. Philipp. J. Sci. 1987, 116, 403; Banerji, J. et al. Indian J. Chem. 1988; 27B, 597; Achari, B. et al. Phytochemistry. 1984, 23, 703]. Stem-bark afforded five new flavone glycosides along with luteolin and acerosin. The new flavone glycosides are 6p-glucopyranosyl-7-hydroxy-3′,4′,5′,8-tetramethoxyflavone-5-O-α-L-rhamnopyranoside, 3′,7-dihydroxy-4′,6,8-trimethoxyflavone-5-O-(6″-O-acetyl-p-D-glucopyranoside),3,3′,4′,6,7,-pentamethoxyflavone 5′-O-(4″-O-β-D-glucopyranosyl-cc-rhamnopyranoside, 4′,5,7-tri-hydroxyflavone-8-(2″-caffeoyl-a-glucopyranoside) and 3′,5,5′,7-tetrahydroxy-4-methoxyflavone -3′-O-(4″-O-α-D-galactopyranosyl) galactopyranoside [Rao, V. K. et al. Indian J. Pharm. 1977,39, 41; Subramamian, P. M. and Misra, G. S. Indian J. Chem. 1978, 16B, 615; Subramaniam, P. M. and Misra, G. S. J. Nat. Prod. 1979,42, 540]. A diterpenoid, 5β-hydro-8,11,13-abieta-trien-6a-ol and three triterpenoids, 2 cc, 3a-dihydroxyoleana-5 12-dien-28-oic acid, 2a, 3a-diacetoxyoleana-5, 12-dien-28-oic acid, 2,3 a-diacetoxy-18-hydroxyoleana-5,12-dien-28-oib acid have been isolated from the seeds, These compounds exhibited antiinflammatory activity [Chawla, A. S., Sharma, A. K., Handa, S. S. and Dhar, K. L. Indian J. Chem. 1991, 30B, 773 and J. Nat. Prod. 1992,55,163. From the roots acetyloleanolic acid was isolated [Vishnol, S. P., Shoeb, A., Kapil, R. S. and Popli, S. P. Phytochemistry 1983, 22, 597].
Five Iridoid glycosides have been reported from the leaves of V. negundo. These areaucubin, agnuside (Hansal et al. Phytochemistry 4, 1965, 9) negundoside, 6′-hydroxyhenzoyl mussaenosidic acid (Sehgal et al. Phytochemistry, 21, 1982, 363) and nishindaside (Datta et al. Tetrahedron 39, 1983, 3067).
Liver disorders are still the major hazard both in urban and rural population. Despite scientific advances in our understandings in the management of liver disorders and the leads provided by traditional system of medicine, no specific treatment of liver ailments is available except chemically undefined a few herbal formulations [Subeamonium and Puspagandan, Indian Journal of Pharmacology, 31, 166-175 (199)]. During our search for hepatoprotective agents of plant origin, the aqueous alcoholic extract of V. negundo and a fraction isolated from its exhibited strong hepatoprotective and immunostimulating activities. A process has been developed for isolation of fraction possessing immunostimulating activity from the leaves of V. negundo for which a patent has been granted to Regional Research Laboratory, Jammu [Sun, J. L. et. al Indian Patent No. 178388 dt. 19-03-97). Another patent application has been submitted by Regional Research Laboratory, Jammu for a process for isolation of a bioactive composition possessing hepatoprotective and immuno-stimulating activity (Application no. 116/DEL98 dt. 16.1.98). iridoid glycosides of Picrorhiza kurroa exhibit strong hepatoprotective activity {Ansari, R. A. et al., Indian J. Med. Research 1988, 87, 401). V. negundo has also been found to contain a number of iridoid glycosides. An iridoid glycosides. 2′-p-hydroxybenzoylmussaenosidic acid (negundoside) was isolated and evaluated for its hepatoprotective activity along with the alcoholic extract of the plant. Both alcoholic extract (coded as 033) and negundoside (coded as 033 (2)) showed marked hepatoprotective activity in experimentally induced hepatic damage with CCl4 and galactosamine (GaIN) in rats. A comparison with the known hepatoprotective agent silymarin revealed that 033 and 033 (2) exhibited higher hepatoprotective potential in most of the parameters with respect to their effect on elevated levels of serum and liver homogenate parameters (Table 1 and 2). Thus the main objective of the present invention is to provide hepatoprotective activity of a bioactive molecule isolated from leaves of V. negundo viz., negundoside of formula 1. As shown in the diagram accompanying this specifications
Accordingly, the present invention provides hepatoprotective activity of a compound of formula 1 accompanying the specifications which comprises    (a) powdering the plant material by known methods;    (b) preparing the aqueous alcoholic extract by percolation;    (c) concentrating the alcoholic extract by conventional method,    (d) removing fatty non polar constituents by triturating the extract with solvents such as ethylene chloride, methylene chloride, chloroform or ethyl acetate;    (e) adsorbing the residue extract over silica gel;    (f) isolation of agnuside from the adsorbed extract by column chromatography;    (g) hepatoprotective activityCharacterisation of Negundoside 1
1 obtained as crystalline compound, mp 148-50° C. M+: 496, UV-260 nm (MeOH) nm, (KBr) spectrum showed absorptions at 3400, 1700, 1640 and 1610 cm′1. ′H NMR (200 MHz, CD3OD) 5 5.48 (J 3, H-1) 7.09 (s, H-3) 25 (m-H-5) 2.19 (m, H-9) 1.25 (s, H-10) 6.80 (dd, 2, 7, H-3″, H-5″), 7.83 (d, 2,7, H-2″, 6″) 4.99 (d,7, H-1′) 4.69 (d, J, 7, H-2″) 13C NMR, 122.99 (C-1), 133.67, (C-2″,6″) 116.92 (C-3″,5″), 164.11 (C-4″) 168.08 (CO) (95.83 (C-1), 151.98 (C-3), 116.92 (C-4), 30.96 (C-5) 31.98 (C-6) 42.04 (C-7), 80.64 (C-8). 53.19 (C-9) 25.15 (C-10), 170.71 (C-11) 98.64 (C-1′), 76-85 (C-2′) 75.70 (C-3′), 72.55 (C-4) 79.30 (C-5′) 63.54 (C-6′).